The BEAF-32 Protein Directly Interacts with Z4/putzig and Chriz/Chromator Proteins in Drosophila melanogaster

Doklady Biochemistry and Biophysics - In Drosophila, the BEAF-32, Z4/putzig, and Chriz/Chromator proteins colocalize in the interbands of polytene chromosomes. It was assumed that these proteins...     

Banding Pattern of Polytene Chromosomes as a Representation of Universal Principles of Chromatin Organization into Topological Domains

Drosophila polytene chromosomes are widely used as a model of eukaryotic interphase chromosomes. The most noticeable feature of polytene chromosome is transverse banding associated with alternation of dense stripes (dark or black bands) and light diffuse areas that encompass alternating less compact gray bands and interbands visible with an electron microscope. In recent years, several approaches have been developed to predict location of morphological structures of polytene chromosomes based on the distribution of proteins on the molecular map of Drosophila genome. Comparison of these structures with the results of analysis of the three-dimensional chromatin organization by the Hi-C method indicates that the morphology of polytene chromosomes represents direct visualization of the interphase nucleus spatial organization into topological domains. Compact black bands correspond to the extended topological domains of inactive chromatin, while interbands are the barriers between the adjacent domains. Here, we discuss the prospects of using polytene chromosomes to study mechanisms of spatial organization of interphase chromosomes, as well as their dynamics and evolution.     

The Chriz–Z4 complex recruits JIL-1 to polytene chromosomes,a requirement for interband-specific phosphorylation of H3S10

The conserved band-interband pattern is thought to reflect the looped-domain organization of insect polytene chromosomes. Previously, we have shown that the chromodomain protein Chriz and the zinc-finger protein Z4 are essentially required for the maintenance of polytene chromosome structure. Here we show that both proteins form a complex that recruits the JIL-1 kinase to polytene chromosomes, enabling local H3S10 phosphorylation of interband nucleosomal histones. Interband targeting domains were identified at the N-terminal regions of Chriz and Z4, and our data suggest partial cooperation of the complex with the BEAF boundary element protein in polytene and diploid cells. Reducing the core component Chriz by RNAi results in destabilization of the complex and a strong reduction of interband-specific histone H3S10 phosphorylation.     

Banding patterns in Drosophila melanogaster polytene chromosomes correlate with DNA-binding protein occupancy

The most enigmatic feature of polytene chromosomes is their banding pattern, the genetic organization of which has been a very attractive puzzle for many years. Recent genome-wide protein mapping efforts have produced a wealth of data for the chromosome proteins of Drosophila cells. Based on their specific protein composition, the chromosomes comprise two types of bands, as well as interbands. These differ in terms of time of replication and specific types of proteins. The interbands are characterized by their association with "active" chromatin proteins, nucleosome remodeling, and origin recognition complexes, and so they have three functions: acting as binding sites for RNA pol II, initiation of replication and nucleosome remodeling of short fragments of DNA. The borders and organization of the same band and interband regions are largely identical, irrespective of the cell type studied. This demonstrates that the banding pattern is a universal principle of the organization of interphase polytene and non-polytene chromosomes.     

Identical functional organization of nonpolytene and polytene chromosomes in Drosophila melanogaster

Salivary gland polytene chromosomes demonstrate banding pattern, genetic meaning of which is an enigma for decades. Till now it is not known how to mark the band/interband borders on physical map of DNA and structures of polytene chromosomes are not characterized in molecular and genetic terms. It is not known either similar banding pattern exists in chromosomes of regular diploid mitotically dividing nonpolytene cells. Using the newly developed approach permitting to identify the interband material and localization data of interband-specific proteins from modENCODE and other genome-wide projects, we identify physical limits of bands and interbands in small cytological region 9F13-10B3 of the X chromosome in D. melanogaster, as well as characterize their general molecular features. Our results suggests that the polytene and interphase cell line chromosomes have practically the same patterns of bands and interbands reflecting, probably, the basic principle of interphase chromosome organization. Two types of bands have been described in chromosomes, early and late-replicating, which differ in many aspects of their protein and genetic content. As appeared, origin recognition complexes are located almost totally in the interbands of chromosomes.     

Chromatin insulators specifically associate with different levels of higher-order chromatin organization in Drosophila

Chromatin insulators are required for proper temporal and spatial expression of genes in metazoans. Here, we have analyzed the distribution of insulator proteins on the 56F–58A region of chromosome 2R in Drosophila polytene chromosomes to assess the role of chromatin insulators in shaping genome architecture. Data show that the suppressor of Hairy-wing protein [Su(Hw)] is found in three structures differentially associated with insulator proteins: bands, interbands, and multi-gene domains of coexpressed genes. Results show that bands are generally formed by condensation of chromatin that belongs to genes containing one or more Su(Hw) binding sites, whereas, in interbands, Su(Hw) sites appear associated with open chromatin. In addition, clusters of coexpressed genes in this region form bands characterized by the lack of CP190 and BEAF-32 insulator proteins. This pattern correlates with the distribution of specific chromatin marks and is conserved in nurse cells, suggesting that this organization may not be limited to one cell type but represents the basic organization of interphasic chromosomes.     

Faint gray bands in Drosophila melanogaster polytene chromosomes are formed by coding sequences of housekeeping genes

Chromosoma - In Drosophila melanogaster, the chromatin of interphase polytene chromosomes appears as alternating decondensed interbands and dense black or thin gray bands. Recently, we uncovered...     

Repetitive arrays containing a housekeeping gene have altered polytene chromosome morphology in Drosophila

Much of our understanding of gene and chromatin organization has been developed from observation of polytene chromosomes. We describe an experimental approach using transgenes that has allowed us to observe local changes in polytene morphology. A composite P transposon that contains a fusion between the regulatory region of Prat, a purine synthesis gene, and brown (bw), an eye pigment reporter, was transformed into the 65A10 polytene band and subjected to P-transposase mutagenesis. Arrays of up to 320 kb at 65A10 were recovered by selection for increased pigment, and pigment levels were found to be proportional to numbers of copies. In polytene chromosomes, the original transformant was found to split 65A10 into two thin bands separated by an interband. With increases in copy number, the interband became progressively denser, eventually forming a dark, amorphous, deformable structure unlike any previously reported. The persistence of Prat expression in development, together with the cytological appearance of these large arrays, suggest that the state of the Prat promoter is affecting polytene structure. Because this unique structure is distinct from bands, interbands, puffs, and the chromocenter, which comprise polytene chromosomes, we suggest that it is composed of an altered form of chromatin.     

The proteins of polytene chromosomes of Drosophila hydei

It is reported that chromatin can be prepared from highly purified polytene nuclei from the salivary glands of third instar larvae of Drosophila hydei; such chromatin differs from that of diploid nuclei mainly by deficiencies in certain nonhistone chromosomal proteins. It is suggested that these proteins are important components of constitutive heterochromatin, which is severely underrepresented in polytene chromosomes. Chromosome morphology, including the pattern of induced puffs, is maintained throughout the mass isolation of glands and sucrose gradient purification of nuclei, as indicated by studies on temperature-shocked and control larvae. No significant alteration in the chromosomal proteins following puff induction by heat shock could be detected on analysis of the isolated protein fractions by disc gel electrophoresis. More sensitive techniques must be developed to study the apparent rearrangement or accumulation of protein at puff sites, and to elucidate the role of this protein in gene activation.     

The location of 5S RNA genes in lampbrush polytene chromosomes from Drosophila

Drosophila polytene chromosomes were transformed into lampbrush-like structures by exposure to solutions of alkali-urea. In this process, the chromosomes shorten and widen, and the bands (chromomeres) extend laterally into loops leaving a central core between the paired homologues. The expanded polytene chromosomes are very similar in appearance to the true lampbrush chromosomes of amphibian oocytes and to ordinary chromosomes in pachytene. The denaturing effects of alkali-urea were partially counteracted by return of the treated chromosomes to Ringer solution. These observations are interpreted in terms of recent findings on protein backbones in chromosomes, and indicate that chromosomes generally may have very similar basic organization, despite differences due to species, polyteny and degree of condensation. To gain more information on the specific location of a structural gene, ¹²⁵I-labelled low molecular weight (containing 5S RNA) was hybridized in situ to normal and lampbrush-like polytene chromosomes. Autoradiography showed silver grain distribution for 5S RNA consistent with hybridization primarily to the loop regions of the lampbrush chromosomes rather than the core. This provides further indirect evidence that structural genes like 5S RNA may be located on the bands (chromomeres) and not the interbands of normal polytene chromosomes.